Professor Nikolay Dokholyan - Biochemistry and Biophysics - UNC School of Medicine

Abstract
We develop new optogenetic and chemogenetic tools to establish control of proteins and signaling cascades directly in living cells for direct interrogation of cellular networks, protein-protein interactions, and the roles of individual proteins in cellular life. To circumvent a fundamental difficulty of establishing such control, which is to go unnoticed by endogenous interaction partners of the target protein, we utilize allosteric sites that are coupled to the active sites. We install small “handles” into determined allosteric sites of the target protein that modulate its activity without affecting endogenous interactions and function. To modulate protein activity with light or a ligand, we utilize small naturally occurring LOV2 for light and artificially designed uniRapR for ligand, correspondingly. Upon irradiation or without the ligand, LOV2 or UniRapR are unstructured; without light or with the ligand, these domains are structured. Through allostery the structural order/disorder is coupled to the active site of the protein rendering it active/inactive. Hence, by light or a ligand we can access the activity of the protein through “invisible” handles installed at a distance from the active site. We demonstrate the utility of the chemogenetic and optogenetic approaches to protein regulation in a number of applications relevant to cellular motility.